The goal of the proposed research is to understand the processes which control differentiation of myogenic cells at a cellular and molecular level. The method of approach will be to analyze various properties of a permanent rat myogenic cell line which exhibits a characteristic muscle development when cultured in vitro under appropriate conditions. By comparing the control of differentiation of this permanent cell line and its subclones with the differentiation of normal diploid muscle cells we hope to elucidate whether cell transformation alters these processes. We will study what controls are exerted in these cells on the expression of the major contractile protein, myosin heavy chain (MHC), during differentiation. We will determine the transcription, processing, and translation of MHC mRNA. The properties of MHC mRNA sequences present in the nucleus and cytoplasm will be examine. MHC mRNA sequences will be assayed by two methods of DNA-RNA hybridization using a cDNA copy (cDNA) of MHC mRNA synthesized with AMV reverse transcriptase. Studies with whole cells will be complemented by experiments designed to study the transcription, processing and transport of MHC mRNA sequences in in vitro isolated nuclei. Using similar techniques we will determine the level of control of ribosome accumulation in growing and differentiated myogenic cells. The other major area of investigation involves the use of cellular genetics for analyzing the regulatory processes controlling muscle differentiation. Using different culture conditions and drugs which will affect differentiation, we will study: a) the kinetics of commitment of myogenic cells to terminal differentiation; b) whether the commitment process is expressed at the level of MHC genes. A second approach will be to study mutant myogenic cells which have an altered differentiation program. Most of the mutants have already been isolated. The mutants will be analyzed biochemically for the presence of several biochemical markers characteristic of muscle differentiation and studied further by complementation analysis using cell hybrids formed between mutants and parent cell lines and between mutants. The studies are relevant to the understanding of heart hypertrophy.